Taq DNA Polymerase is supplied with the unique QIAGEN PCR Buffer that minimizes the need for optimization of PCR parameters, as well as Q-Solution, a novel additive that enables efficient amplification of "difficult" (e.g., GC rich) templates. In addition, CoralLoad PCR Buffer (containing two gel-tracking dyes) is also provided, enabling immediate loading of PCR products.

[A] CoralLoad Concentrate contains two gel-tracking dyes,enabling immediate gel loading of PCR products for [B ] easy visualization of DNA migration.|PCR amplification at the indicated Mg2+ concentrations using QIAGEN PCR Buffer and Taq DNA Polymerase (QIAGEN). The same PCR was performed in parallel using a PCR buffer and Taq polymerase from another supplier (Supplier AII). The single-copy human prion protein gene was amplified successfully in each case using the buffer and enzyme from QIAGEN. M: markers.|Two different primer-template systems were amplified in duplicate using QIAGEN PCR Buffer and Taq DNA Polymerase in the absence (–) or presence (+) of 1x Q-Solution. Q-Solution enables specific amplification of difficult templates. [A] human angiotensin receptor II gene; [B] mouse protein kinase C gene; M: markers.|The human single-copy cystic fibrosis gene was amplified with Taq DNA Polymerase and QIAGEN PCR Buffer using the indicated annealing temperatures. Primers employed were a 22mer with a Tm of 57.5°C (GC content: 54.5%) and a 32mer with a Tm of 85.2°C (GC content: 78%). For analysis, 10% of a 100 µl reaction was loaded. M: markers.|A fragment of the single-copy gene for cystic fibrosis was amplified from 30 ng, 3 ng, and 300 pg human genomic DNA corresponding to 104, 103, and 102 copies of target template, respectively. Three different lots of Taq DNA Polymerase were used and equal volumes of the PCR product were analyzed on a 1% agarose gel. M: markers.|Three different-sized products from human genomic DNA were amplified using either Taq DNA Polymerase and QIAGEN PCR Buffer (QIAGEN), or Taq polymerase and a buffer from another supplier (Supplier AII). For analysis, 10% of each reaction was loaded on the gel. Results from duplicate PCR amplifications are shown. M: markers.|Ammonium and potassium cations in QIAGEN PCR Buffers increase specificity of primer annealing. K+ binds to the phosphate groups (P–) on the DNA backbone, stabilizing the annealing of the primers to the template. NH4+, which exists both as the ammonium ion and as ammonia under thermal-cycling conditions, can interact with the hydrogen bonds between the bases (B), destabilizing the weak hydrogen bonds at mismatched bases. The combined effect of the two cations maintains the high ratio of specific-to-nonspecific primer-template binding over a wide temperature range.|

Taq DNA Polymerase outperformed kits tested from other suppliers and delivers robust PCR performance in a wide range of PCR conditions, without the need for time-consuming optimization (see figures "Tolerance of different primer Tm Values" and "Specific amplification of long PCR products"). Every lot of TaqDNA Polymerase is subjected to a comprehensive range of quality control tests, including a stringent PCR specificity and reproducibility assay in which low-copy targets are amplified from human genomic DNA (see figure "Lot-to-lot reproducibility"). The unique formulation of QIAGEN PCR Buffer and CoralLoad PCR Buffer, also provided with the kit, enable highly specific PCR in a variety of PCR conditions with minimal optimization requirements (see figures "Wide annealing-temperature window" and "Tolerance to variable magnesium concentration"). In addition, CoralLoad PCR Buffer enables immediate loading of PCR products onto an agarose gel for even easier handling and faster results. Suboptimal PCR can be improved using Q-Solution, a PCR additive, also provided with the kit (see figure "Amplification of difficult templates").

QIAGEN PCR Buffer

Innovative QIAGEN PCR Buffer has been developed to save time and effort by reducing the need for PCR optimization. QIAGEN PCR Buffer contains both KCl and (NH4)2SO4(see figure "Increased specificity of primer annealing"). This unique buffer facilitates the amplification of specific PCR products. During the annealing step of every PCR cycle, the buffer allows a high ratio of specific-to-nonspecific primer binding. Owing to a uniquely balanced combination of KCl and (NH4)2SO4, the PCR buffer provides stringent primer-annealing conditions over a wider range of annealing temperatures and Mg2+ concentrations than conventional PCR buffers. Optimization of PCR by varying the annealing temperature or the Mg2+ concentration is dramatically reduced and often not required (see figures "Wide annealing temperature window" and "Tolerance to variable magnesium concentration").

CoralLoad PCR Buffer

CoralLoad PCR Buffer has all the advantages of QIAGEN PCR Buffer. In addition, it can also be used to directly load the PCR reaction onto an agarose gel — separate addition of a gel loading buffer is not required. CoralLoad PCR Buffer provides the same high PCR specificity and minimal reaction optimization as the conventional QIAGEN PCR Buffer. Additionally, it contains two marker dyes — an orange dye and a red dye — that facilitate estimation of DNA migration distance and optimization of agarose gel run time (see figure "CoralLoad PCR Buffer"). The buffer ensures improved pipetting visibility and enables direct loading of PCR products onto a gel, for enhanced convenience.

Q-Solution

Q-Solution facilitates amplification of GC-rich templates or templates with a high degree of secondary structure by modifying the melting behavior of DNA. Use of this unique reagent often enables or improves suboptimal PCR (see figure "Amplification of difficult templates"). Unlike DMSO and other PCR additives, Q-Solution is used at a defined working concentration with any primer–template system and is not toxic.

Procedure

Taq DNA Polymerase ensures highly specific PCR for a range of applications — with minimal optimization of PCR parameters. The streamlined, easy-to-follow protocol provided with the kit simplifies PCR setup. For added convenience and easier handling, CoralLoad PCR Buffer is provided. PCR products can be directly loaded onto a gel without the addition of a loading dye. To ensure success with GC-rich templates, the PCR enhancer Q-Solution is included.

Applications

Taq DNA Polymerase is used for standard and specialized applications, including: